Mitrani-Rosenbaum S, Maroteaux L, Mory Y, Revel M, Howley P M
Mol Cell Biol. 1983 Feb;3(2):233-40. doi: 10.1128/mcb.3.2.233-240.1983.
A 1.6-kilobase DNA segment of the genomic human interferon beta 1 (IF-beta 1) gene was inserted into each of two possible orientations at the single HindIII site of a recombinant plasmid pBPV69T, consisting of the 69% transforming region of the bovine papilloma virus type 1 (BPV-1) and a modified SalI-SalI fragment of plasmid pBR322. After cleavage of the pBR322 sequences from this recombinant, BPV69T-IF-beta 1 hybrid DNAs were transfected onto C127 mouse cells by the standard calcium precipitation technique. Mouse cells transformed by this hybrid DNA produced low levels of human IF-beta 1 constitutively and responded to induction with either inactivated Newcastle disease virus or polyriboinosinic acid-polyribocytidylic acid. The BPV69T-IF-beta 1 hybrid DNA was nonintegrated in the transformed mouse cells but had acquired DNA sequences as a result of the transfection. Accurate transcripts of the IF-beta 1 mRNA were detected in cells only after induction. When the IF-beta 1 gene was oriented in the plasmid in the same direction of transcription as the BPV-1 genome, transcription was promoted from within the BPV-1 sequences. These results indicate that the regulatory sequences responsible for the inducible expression of the human IF-beta 1 gene are present in the 1.6-kilobase genomic segment and that these sequences can function in a free extrachromosomal state linked to BPV-1 sequences.
将基因组人干扰素β1（IF-β1）基因的一个1.6千碱基的DNA片段以两种可能的方向插入到重组质粒pBPV69T的单一HindIII位点，该重组质粒由1型牛乳头瘤病毒（BPV-1）的69%转化区域和质粒pBR322的一个修饰的SalI-SalI片段组成。从该重组体中切割掉pBR322序列后，通过标准的钙沉淀技术将BPV69T-IF-β1杂交DNA转染到C127小鼠细胞上。由这种杂交DNA转化的小鼠细胞组成性地产生低水平的人IF-β1，并对用灭活的新城疫病毒或聚肌苷酸-聚胞苷酸诱导产生反应。BPV69T-IF-β1杂交DNA在转化的小鼠细胞中未整合，但由于转染而获得了DNA序列。仅在诱导后才在细胞中检测到IF-β1 mRNA的准确转录本。当IF-β1基因在质粒中的方向与BPV-1基因组的转录方向相同时，转录从BPV-1序列内部被促进。这些结果表明，负责诱导性表达人IF-β1基因的调控序列存在于1.6千碱基的基因组片段中，并且这些序列可以在与BPV-1序列相连的游离染色体外状态下起作用。
